The macromolecular x-ray diffraction facility of the University of Colorado, Boulder proposes to upgrade an X-ray source: specifically, a Rigaku RUH2R rotating anode X-ray generator with Osmic "blue" confocal multilayer mirrors will be upgraded to a Rigaku MM007 generator with Varimax HR (="high resolution") confocal mirrors. We are receiving a used MM007 as a gift, and request funds only for the confocal mirrors. The x-ray facility is currently used by three major and seven minor NIH-supported users. Crystallographic projects currently underway include: large catalytic RNAs (Cech and Pace) and riboswitches (Batey), RNA- protein and DNA-protein complexes such as those found in the telomerase system (Cech and Wuttke) and specific recognition complexes (McKay), molecular chaperone and integral membrane proteins (Sousa), and several RNAs and proteins involved in specific biological processes (Ahn, Copley, Pardi, Falke). Many of these projects pose significant challenges in their early stages;typically, small preliminary crystals from a large number of constructs and/or crystallization conditions must be screened for diffraction to determine whether they are worth pursuing. Additional challenges often present themselves during screens for cryoprotectants or suitable phasing derivatives. Finally, in many cases, complete datasets are collected in- house, either prior to or as an alternative to synchrotron data collection. The requested upgrade will enhance the brightness of our X-ray source severalfold (approximately sixfold according to Rigaku's data), thereby enhancing the crystal screening and data collection capability of our in-house facilities. PUBLIC HEALTH RELEVANCE: The three-dimensional structures of biologically significant macromolecules provide the foundation for modern molecular medicine and rational drug design. Structural studies at the University of Colorado, Boulder encompass ribozymes (catalytic RNAs), telomerase, membrane protein structure and folding, and bacterial drug resistance, to name a few areas. The shared X-ray crystallography facility, for which instrumentation is requested, provides resources (both instrumental and intellectual) to a broad spectrum of users who pursue structural studies on the campus.